Fibroblasts are the principal cells that reside in the dermis of the skin and stroma of all vertebrate organs. Although fibroblasts were thought by many to be homogenous, we have discovered that human fibroblasts from different anatomic sites of the skin and internal organs constitute many distinct, differentiated cell types that carry out unique genetic programs to specify the site-specific design and functions of epithelial organs. The broad goal of this project is to understand the physiologic specialization and developmental regulation of fibroblasts by following the expression level of a large number of genes in cultured fibroblasts. Firstly, using high density cDNA microarrays and computational bioinformatics, we will create a comprehensive molecular map of fibroblast gene expression patterns from diverse site in the human body. This large compendium of fibroblast gene expression patterns will reveal how many different types of fibroblasts exist, how they are related to one another, and where they are located in the human body. The gene expression programs that characterize specific fibroblasts will define the physiologic specialization of different fibroblast cell types and will help to explain the inductive potentials of fibroblasts in development, their functions during tissue repair, and the anatomic specificity of diseases affecting skin and connective tissues. Secondly, the site-specific gene expression profiles of fibroblasts will be used as tools to understand how different types fibroblasts develop. By assessing the gene expression patterns of fibroblasts from different sites cultured together and of fibroblasts cultured in isolation from one another, we will address whether fibroblasts instruct each other to adopt site-specific fates, or whether the differentiation program is controlled autonomously within each cell. Thirdly, adult fibroblasts have the unique property of maintaining positional memory, preserved in the form of Hox gene expression patterns established during embryogenesis. Hox genes encode a family of transcription factors that activate other genes to specify positional identities in development. We will test whether Hox genes are master regulators of fibroblast differentiation by assessing the gene expression patterns of fibroblasts with ectopic or inactivated Hox functions. From the proposed experiments, we hope to learn a great deal about the gene expression programs in fibroblast differentiation. This knowledge will provide many basic insights into wound healing, organ development, anatomic specificity of genetic and acquired diseases affecting the skin and connective tissues; and shed light on how positional identity is acquired and maintained in the human body.